Prior studies demonstrated that histotripsy generated by high-intensity tone bursts to excite a bubble cloud adjacent to a medical implant can destroy the bacteria biofilm responsible for the infection. The goal of this study was to treat S. aureus biofilms on surgical mesh samples while varying the number of cycles in the tone burst to minimize collateral tissue damage while maximizing therapy effectiveness. S. aureus biofilms were grown on 1 cm square surgical mesh samples. The biofilms were then treated in vitro using a spherically focused transducer (1.1 MHz, 12.9 cm focal length, 12.7 cm diameter) using either a sham exposure or histotripsy pulses with tone burst durations of 3, 5, or 10 cycles (pulse repetition frequency of 333 Hz, peak compressional pressure of 150 MPa, peak rarefactional pressure of 17 MPa). After treatment, the number of colony forming units (CFUs) on the mesh and the surrounding gel was independently determined. The number of CFUs remaining on the mesh for the sham exposure (4.8±0.9-log10) (sample mean ± sample standard deviation-log10 from 15 observations) was statistically significantly different from the 3-cycle (1.9±1.5-log10), 5-cycle (2.2±1.1-log10), and 10-cycle exposures (1.0±1.5-log10) with an average reduction in the number of CFUs of 3.1-log10. The numbers of CFUs released into the gel for both the sham and exposure groups were the same within a bound of 0.86-log10, but this interval was too large to deduce the fate of the bacteria in the biofilm following the treatment.